An important aspect of immunotherapy or cell therapy utilizes modified antigen-presenting cells (APC) to elicit an immune response in patients. Dendritic cells (DC), the most potent APCs, are essential for the induction and maintenance of this response and are central to the development of effective immunotherapies towards treating animal and human infectious diseases as well as different human cancers. For example, in tumor tissues, DCs can engulf apoptotic or necrotic tumor cells, process and present tumor antigens to induce tumor specific T cell responses and immunity. However, despite this potent defensive barrier, tumors progress, metastasize, and eventually can kill the host. Immunosuppressive factors secreted in the tumor microenvironment contribute to the evasion of immune surveillance. Such factors can profoundly affect DC function. Strong evidence indicates that DC defects in cancer are due to abnormal differentiation and maturation of myeloid cells, which has deleterious effects on T cell activation and antitumor response. Therefore, the efficacy of DC-based immunotherapies is compromised by immunosuppressive tumor microenvironments. Cancer patients display a significant reduction of mature and functional DCs, as well as accumulation of immature DCs (iDCs) or immature myeloid cells (iMCs, DC progenitors from the bone marrow), which has been associated with increased plasma levels of suppressive tumor-derived factors. DCs derived from cancer patients express no or low levels of costimulatory molecules CD80 and CD86. Consequently, the major hurdle in DC immunotherapy is that iDCs or iMCs induce T cell tolerance or anergy, which consistently affects the development of an effective anti-tumor response while favouring tumor growth and metastasis.
Dysregulation of one of the critical pathways essential for the activation of the immune response, the cytokine-activated janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway has been identified as one of the key factors responsible for the abnormal DC differentiation and function in cancer. This pathway controls DC differentiation, maturation, activation and DC-dependent induction of TH1-cell differentiation. There are four members in the Jak family of tyrosine kinases: Jak1, Jak2, Jak3, and Tyk2 [1,2]. Jak kinases are constitutively associated with cytokine receptors and are activated by the binding of a cytokine to its cell-surface receptor. Once activated the Jak kinase phosphorylates specific tyrosine residues on the receptor providing binding sites for STATs. STATs, consisting of 7 members: Stat1, Stat2, Stat3, Stat4, Stat5a, Stat5b, and Stat6, are a group of latent cytoplasmic transcription factors, which reside in an inactive form in the cytoplasm. They are activated by binding to the phosphorylation sites on cytokine receptors and are subsequently phosphorylated on a specific tyrosine residue by Jak. When phosphorylated they disassociate from the receptor, dimerize and enter the nucleus to induce expression of target genes. In addition to cytokine and chemokine receptors, JAK-STAT signaling can also be initiated directly by receptor tyrosine kinases such as Epidermal growth factor receptor, platelet derived growth factor receptor and others. It is the combination of these different stimuli, positive and negative; as well as the recognition of immunogens that lead to optimal potentiation of DCs and the downstream activation of Th-1 cell differentiation.
Currently, the generation of DCs for immunotherapy treatment, by in vitro differentiation of monocytes from peripheral blood and their subsequent maturation and activation, is through the use of various cytokine cocktails. The synergistic interaction between GM-CSF and IL-4 induces JAK-STAT pathway activation during DC differentiation. IL-4 mediates its effects primarily through STAT6 activation and GM-CSF activates STAT1 and STAT5. STAT6 is constitutively activated in immature DCs and declines as the cell differentiates into a functionally mature DC, whereas STAT1 signaling is more robust in mature DCs, correlating with upregulation of costimulatory molecules expression and IL-12 production. STAT5 is required for differentiation of DCs during maturation. One additional STAT required for the early phases of DC differentiation, as well as commitment of common lymphoid progenitors (CLP) and common myeloid progenitors (CMP) to the DC lineage is STAT3. DC differentiation inversely correlates with STAT3 activation hence mature DCs show low levels of STAT3 activation. STAT3 hyperactivation however results in inhibition of DC maturation/activation in response to diverse stimuli. Most of the tumor-derived factors take advantage of this pathway to promote abnormal DC differentiation by inducing STAT3 hyperactivation. Activated STAT3 decreases intracellular major histocompatibility complex II (MHCII) alpha/beta dimers, and H2-DM levels in DCs by increasing cathepsin S activity. In addition, STAT3 hyperactivation also inhibits LPS-induced interleukin (IL)-12p40 gene expression and affects NF-kB recruitment to the IL-12p40 promoter leading to a build-up of functionally impaired and immature myeloid cells with a high immunosuppressive potential. These reports indicate that STAT 3 activation must be tightly controlled in order to maintain the balance between inhibitory and activating signals.
PTPs are a family of transmembrane or intracellular enzymes that control multiple cellular regulatory processes by dephosphorylating phospho-tyrosine substrates. There are 107 PTPs in the human genome and several reviews have described in detail the members of this gene family. TC-PTP (PTPN2) is found principally as a ˜45 kD intracellular protein that localizes primarily to the nucleus and belongs to the class I subfamily of phospho-tyrosine specific PTPs. Although TC-PTP is ubiquitously expressed, highest expression is observed in all hematopoietic cells [1, 3, 4]. In vitro analysis indicates that TC-PTP negatively regulates cytokine signaling by inhibiting JAK-STAT pathways. Currently, JAK1, JAK3, STAT1, STAT3, and STAT5a/b have been identified as putative TC-PTP targets downstream of cytokines such as IL-2, IL-6, IL-4, and IFN-γ [1, 2, 5]. Consequently, TC-PTP has been identified as a critical negative regulator of DC activation, suggesting that inhibitors of this enzyme may be beneficial in the activation of these cells for immunotherapy treatment of disease.